Transmissible spongiform encephalopathies (TSEs or prion disease) are fatal neurodegenerative diseases such as scrapie, Creutzfeldt-Jakob disease (CJD), bovine spongiform encephalopathy and chronic wasting disease (CWD). Our project is aimed at understanding and blocking the conversion of normal PrP (PrP-sen) to PrP-res, the abnormal form of prion protein (PrP) that appears to underlie TSE transmission and pathogenesis. We have employed a wide variety of cell biological, biochemical, biophysical and in vivo experimental approaches. Over the last year we have 1)continued the identification and isolation of the most and smallest infectious units of TSE diseases using field flow fractionation, light scattering, electron microscopy and electophoresis, 2) discovered degenerate phosphorothioate oligonucleotides as potent PrP-res inhibitors and anti-TSE drugs in vitro and in vivo, 3)characterized the process by which PrP-res invades neurons and glial cells, initiates infection, and is transported along neuronal processes to extremities where contact is made with other cells, using confocal microscopy, 4) developed the first tissue culture cell line infected with CWD and identified the first inhibitors of CWD replication, 5) continued to test porphyrins and other inhibitors for prophylactic and therapeutic activities against scrapie in animals, and identified one with efficacy against an established central nervous system infection, 6) continued to adapt chronic wasting disease from deer and elk to hamsters and transgenic mice expressing hamster PrP-sen, 7) probed PrP-res conformation using amino acid derivitization and mass spectrometry, 8) analyzed the effects of PrP mutations on PrP conversion to PrP-res